A variety of plasmid DNA replication mutants in Escherichia coli where the mutation is localized on the host chromosome or on the plasmid DNA molecule are being isolated and characterized in an attempt to delineate the genetic and biochemical factors involved in the controlled duplication of circular DNA plasmid elements. Plasmids examined in this study include colicinogenic factors E1, E2, V (ColE1, ColE2, ColV, respectively), the F1 sex factor and the antibiotic resistance transfer factors R1, R64, R100, R6K and R28K. Emphasis is being placed on the biochemical characterization of relaxation complexes of these various plasmid elements and the role of these complexes in the replication and conjugal transfer of these plasmids. Replicative intermediates of these circular DNA elements are being isolated from intact and mini-cells of E. coli and the structure of these intermediates determined by physical- chemical methods and the use of DNA restriction enzymes. Finally, plasmid DNA elements are being reconstructed in vitro as a means of delineating the genetic regions of plasmid DNA vital to the replication of the plasmid element.